Infectious diseases caused by facultative or strict intracellular microbes or microorganisms, including a significant amount of bacteria, protozoa, fungi and viruses against which no effective vaccine or medication for wide-scale use is available, remain a torment for mankind.
These microorganisms of many species and genera are mostly intracellular parasites in their life-cycle, therefore called facultative intracellular microorganisms or else strict intracellular microorganisms when their life-cycle occurs entirely inside the host cells, as it is the case of viruses and some pathogenic parasite species.
They are also called opportunistic microorganisms because of their additional ability to survive for long periods inside cells and tissues, including defense cells such as the macrophages that they invade, taking advantage of favorable conditions (e.g. deficient immunity), to easily spread over with devastating and often lethal consequences.
According to estimates, several million people become annually infected with the bacterium or mycobacterium called Mycobacterium tuberculosis (M. tuberculosis) which is a typical intracellular microorganism, resulting in more than one million deaths (Dye C, Scheele S, Dolin P, Pathania V, Raviglione M C. Consensus statement. Global burden of tuberculosis: estimated incidence, prevalence, and mortality by country. WHO global surveillance and monitoring project. (JAMA 1999 Aug. 18; 282(7): 677-86).
Although anti-tuberculosis medications are available, the existing treatments are complex, long and include administration of several substances with antiparasitic properties for at least four months, which considerably decreases patient adherence.
Moreover, the emergence of types of parasites resistant to the current medications and therapies aggravate an already serious public health problem. The low rate of efficacy of medications and treatments combined with the emergence of types of resistant microorganisms deepen the challenge of tuberculosis and other infectious diseases caused by intracellular microorganisms.
Malaria is another infectious disease with worldwide repercussion caused by an intracellular parasite of the Plasmodium genus. It is considered the world's most important parasitic disease. It is endemic in 91 countries and an estimated 300 to 500 million malaria cases occur each year. The most severe forms of malaria occur mostly in tropical regions. It is the seventh leading cause of death, accounting for around 2 million annual deaths and its control remains a high priority. (The World Health Report 1996, WHO, Geneva, 1997) [[e]] and (World malaria situation in 1994. Wkly Epidemiol Rec 1997; 36:269-75; 37:277-83; 38:285-90).
Besides the use of purely prophylactic measures e.g. killing the vector insects, malaria can be treated using several current or proposed medications and procedures ranging from the use of substances with antiparasitic properties (antimalarials), immunization or vaccination, to the use of immunotherapy that in the current state of the art is based on the administration of some types of cytokines, particularly interferons, to the infected patients.
The main problems in malaria control, concerning the prophylactic measures, are the emergence of new types of vectors of the disease, particularly the mosquitoes of the Anopheles genus which become increasingly resistant to insecticides, as well as severe environmental problems associated to the indiscriminate use of insecticides.
The success of the treatment of various forms of malaria has been limited by the undesirable side effects of current antimalarial drugs, and also due to restrictions on taking these medications during pregnancy (with concomitant infection), dissemination of varieties of resistant protozoa and, finally, the inexistence of highly effective vaccines.
Leprosy or hanseniasis is another infectious disease caused by intracellular microorganisms, the Mycobacterium leprae (M. leprae) that belongs to the same genus as M. tuberculosis. The M. leprae causes skin lesions and severe nerve damage. It is endemic in some tropical countries, particularly in Asia, and despite advancements in the treatment, the disease is far from being under control, affecting large populations, according to data from the World Health Organization. (World Health Organization Global Strategy for further reducing the leprosy burden and sustaining leprosy control activities: plan period: 2006-2010. (Geneva (SWT): WHO; 2005).
The M. leprae, like the M. tuberculosis, multiplies very slowly compared to other bacteria. The immune system is often able to provide an appropriate response to mycobacteria, which results in the formation of granulomas. The type of reaction of the immune system to the M. leprae plays a very important role in the progression of the disease, since this mycobacterium also survives phagocytosis and is therefore able to multiply inside the macrophage, like the M. tuberculosis. 
In the case of inflammatory reaction of the immune system, with granuloma formation and the destruction of infected macrophages, the disease turns out to be almost benign because the progression rate is considerably reduced. It is called tuberculoid leprosy.
If no inflammatory reaction occurs, there is no granuloma formation and the bacteria may become rapidly disseminated causing the typical leprosy or lepromatous leprosy. The treatment is based on a combination antibiotics and sulfas aimed at maximizing efficacy and preventing the emergence of varieties resistant to treatment. The World Health Organization (WHO) recommends the association of three drugs: Dapsone (diaminodiphenylsulfone), Rifampicin (an antibiotic of the rifamide group) and Clofazimine (a rimino-phenazine liposoluble dye).
The typical treatment period of leprosy patients varies between 6 and 24 months. The success of this treatment has been limited by its length and the occurrence of major side effects caused by the medications, which considerably decreases patient adherence to the therapy.
Another intracellular microorganism with infection and survival mechanisms similar to those of M. tuberculosis, M. avium and M. leprae is the Listeria monocytogenes (L. monocytogenes) that causes lysteriosis.
Lysteriosis is the name given to a group of clinical symptoms caused by the L. monocytogenes bacterium, including septicemia, meningitis, and encephalitis, cervical or intra-uterine infection. In pregnant women the infection can cause abortion of the phoetus, or premature birth. Other damage may occur such as endocarditis, hepatic lesions and, in other organs, internal or external abscesses and severe skin lesions. The lethality rate of disease is 30% in neonates; 35% in adults; around 11% in patients under 40 and 63% in patients over 60 years old. In the case of septicemia associated to infection by L. monocytogenes the lethality rate is 50% and in the case of meningitis lethality rate can reach 70%.
The recommended treatment, described till the present state of the art and not totally reliable, is based on the use of antibiotics such as penicillins or ampicillins, either alone or in combination with aminoglycosides, such as amikacin. Cephalosporins are not effective. Cases of resistance of L. monocytogenes to antibiotics of the tetracycline group have been recently observed.
Finally, viruses are strict intracellular microorganisms or parasites that cause significant disease in both man and animals. The existing treatments of viral infections in men and animals are mostly based on preventive vaccination, only available for few virus species. The available treatment of viral infections where no vaccine currently exists, or when the host is already infected, described in the state of the art, consists of some combinations of substances of antiviral properties that will be demonstrated in the present report.
A recent development for the clinical treatment of viral infections and some other infectious diseases that has become widespread is based on the use of exogenous cytokines with immunostimulant or immunomodulatory properties, such as the Interferon-alpha (IFN-alpha), Interferon-beta (IFN-beta), Interferon-gamma (IFN-gamma) and Interleukin-2 (IL-2) used in combination or association with other substances (e.g. antiviral substances).
Interferons are glycoproteins of the cytokine family, produced by some cells of the immune system of more complex organisms, mostly T-lymphocytes and a few other specialized cells, in response to external agents that harm the host body, such as viruses, parasites and tumor cells. In the present state of the art, exogenous interferons are obtained by means of cell culture techniques and/or recombinant DNA technologies and some of its derivatives, such as pegylated are obtained by means of chemical semi-synthesis techniques. In the present state of the art, exogenous interferons are used in the treatment of a few types of cancer, as an adjuvant therapy for some infectious diseases, including viral infections and usually associated to antimicrobials, such as antiviral substances and others, as shall be explained in the present report.
A scientific justification provided for the use of the abovementioned combination of substances is the fact that the Interferon has an indirect antimicrobial activity, reinforcing the immune system action by delaying or decreasing the rates of viral replication, thus, improving the success rate of this treatment associated to other therapies. However, the clinical use of exogenous cytokines such as Interferon-alpha, Interferon-beta, Interferon-gamma and Interleukin-2 is plagued by considerable problems, such as the occurrence of noteworthy side effects and high treatment cost. Furthermore, exogenous cytokines used alone in clinical practice as immunomodulators (such as the Interferon-alpha, Interferon-beta, Interferon-gamma and the Interleukin-2) do not show satisfactory activity against viruses and other infectious agents.
In the present state of the art, despite the latest advancements in science and medicine, there is no fully effective treatment (medications or vaccines) against a wide range of intracellular pathogens, particularly viruses, after the occurrence of infection.
The Immune System and its Host Defense Mechanisms Against Intracellular Pathogens—Types of Immune Response.
It is widely known that the immune response plays a fundamental role in the host defense against infectious agents and that it constitutes the main obstacle to widespread occurrence of infections, usually associated with a high mortality rate. It is also known that the number of individuals exposed to infection is much higher than that of individuals affected by the disease, indicating that most living systems, including the human genus, are capable of eliminating these microorganisms and prevent disease progression.
This defense capacity is ensured by the immune system (Janeway C A Jr.—How the immune system protects the host from infection.—Microbes Infect. 2001; 3:1167-71).
Among the various cellular populations that form the immune system, also called immunologic system, are the T4 cells (or T4 lymphocytes, or else T-Helper lymphocytes). The T4 lymphocytes control immune response.
The T4 lymphocytes are responsible for eliminating pathogens, mostly by releasing cytokines and activating or inhibiting all other immune system cells, in order to enable these cells to perform their specific functions of host defense against pathogens.
Types of Immune Response—Main Characteristics.
Acting in host defense, the T4 cells can generate two basic types of immune response known in the state of the art, which are called TH-1 type immune response and TH-2 type immune response.
The TH-1 type immune response, also known as inflammatory response, is characterized by the production of typical cytokines, such as the Interleukin-2 (IL-2), Interferon-gamma (IFN-gamma) and Tumor Necrosis Factors (TNF) by immune system cells. This response activates macrophages, phagocytosis, and finally, the cytotoxic mechanisms of macrophages. The TH-1 type immune response is very effective against strict and facultative intracellular pathogens, such as viruses, protozoa and fungi. It is also effective against various malignant tumors.
The TH-2 type immune response is characterized by the secretion of other cytokines, such as the IL-4 and IL-5 by immune system cells. It also stimulates antibody production by B lymphocytes being effective against organisms that circulate in the bloodstream, or microorganisms such as extracellular bacteria and some parasite species.
The Immune System—Mechanisms and Situations that Favor Pathogens Survival and Infection.
Despite the widely known efficiency with which the immune system has been fighting and controlling invading pathogens, throughout the evolution of mammals, including the human species, several types of facultative or strict intracellular microorganisms have managed to establish survival mechanisms by which they block or reduce the efficiency of the immune response of the host body, particularly the TH-1 type immune response. Unfortunately, this ability to reduce the efficacy of the host immune system, which is a highly specialized form of evolutionary adaptation and survival of microorganisms or infectious agents, can be disastrous to the host, including humans. In fact, the persistence of pathogens in the host body, made possible by inhibition or blockage of the host immune system, may frequently lead to chronic diseases or death. Thus, during the process by which the pathogen enters the host, called acute infection phase or during the process of pathogen persistency in the host organism, called chronic infection phase, the survival of strict or facultative intracellular microorganisms often depends on the occurrence and/or maintenance of a poor immune status of the host body.
This status of defective or deficient immunity, which greatly favors colonization by intracellular pathogens, such as the M. tuberculosis (Mycobacterium tuberculosis), M. leprae (Mycobacterium tuberculosis), M. avium (Mycobacterium avium-intracellulare) and L. monocytogenes (Listeria monocytogenes) is basically related to the decrease or suppression of host's TH-1 type immune response, also called inflammatory response.
This host's defective or suppressed immune response can be either caused by pathogens themselves or be associated to other organic causes and conditions, occurring concomitantly with or prior to infection, which will increase the probability of success of this type of pathogens.
This dysfunction can be directly associated to parasite action by means of several complex mechanisms capable of reducing the endogenous production or release of host cytokines, such as the Interferon-gamma, leading to dysfunction or functional inaptitude of vital components of the host immune system, such as the macrophages, for example, that are cytokine-activated. (J. Gong et al., Infec. and Imm., March 1996, Vol. 64, No. 3. p. 913-918).
The referred condition of immune dysfunction can also be found in other diseases caused by other types of intracellular microorganisms, such as malaria. There is strong evidence in the medical literature supporting an association of depression in TH-1 type immune response, which is related to a significant decrease in endogenous production of Interferon-gamma, to increased disease severity.
Pathologies or situations, such as Diabetes mellitus and Acquired Immunodeficiency Syndrome (AIDS) are other examples of clinical pictures associated with poor immune status, that occurs concomitantly with or prior to infection by pathogens and, as a consequence, facilitate their dissemination in the host body.
The secretion of the cytokines that activate TH-1 type immune response, such as Interleukin-2 (IL-2) and antiviral Interferon (IFN-gamma) by the host body, usually decreases during the HIV infection process. (S. Crowe et al., Antiv. Chemistry & Chem. 12:133-150 Review, 2001), causing severe immunodeficiency mostly due to the progressive suppression of the TH-1 type immune response. This leads to the widespread occurrence of opportunistic infections caused by microorganisms that have been so far successfully controlled by the TH-1 type immune response.
Decrease or virtual suppression of the referred immune response caused by external causes occur often in patients undergoing chemotherapy, radiotherapy or else taking corticosteroids and other medications with immunodepressive properties, which favor the widespread dissemination of intracellular microorganisms.
Based on these facts and information it is possible to affirm that the common aspect of all genera and species of facultative or strict intracellular pathogens, such as bacteria or mycobacteria (M. tuberculosis, M. leprae, M. avium, and L. monocytogenes), protozoa (Plasmodium sp, Leishmania sp), fungi (Candida sp, Cryptosporidium sp) and several classes and types of viruses is their widely known dependence on the existence of a defective immunity condition, also called immune deficiency state of the host organism, that allows and/or facilitates the introduction, survival and multiplication of these pathogens.
Use of Immunomodulators in the Treatment of Intracellular Pathogens—Use of Exogenous Interferon.
Due to the fact that facultative or strict intracellular parasites of many genera and species depend on the occurrence of defective immunity or immune deficiency of host body to successfully enter, survive and multiply, a therapeutic strategy in the current state of the art medicine has been elaborated. It is based on immunotherapy, or the use of a combination of substances with immunomodulatory action or stimulation to revert or minimize immune deficiency in order to provide the host body with the ability to appropriately respond to pathogens.
In order to achieve this purpose, some combinations of substances with immunomodulatory properties, described in the state of the art, are used in clinical practice, as will be demonstrated in more detail in the present report.
In short, this strategy to eliminate intracellular microorganisms involves the use of substances capable of enhancing the immune system, which, in turn, becomes able to appropriately respond to pathogens or in other words: an indirect defense strategy against pathogens.
One of the most widely known substances of this class is the interferons. There are various types of interferons, among which the alpha types (IFN-alpha), beta types (IFN-beta) and gamma types (IFN-gamma), all of them with immunomodulatory properties that are being studied for use in the treatment of several pathologies.
These several types of interferons are being used or considered to be used in the stimulation of the immune system of patients, and are also expected to be used in the treatment of diseases caused by intracellular pathogens, such as tuberculosis.
As it was mentioned before, during the process of HIV-1 (AIDS) infection, the secretion of TH-1 type cytokines, such as Interleukin-2 (IL-2) and antiviral interferon—IFN (gamma) usually decreases (S. Crowe et al., Antiv. Chemistry & Chem. 12:133-150 Review, 2001), leading to the progressive emergence of several types of infections caused by microorganisms that have been so far successfully controlled by the immune system.
Therefore, due to the similarity of mechanisms used by most intracellular parasites to infect host tissues, among them the parasites that cause tuberculosis, or else, their ability to produce or take advantage of host immunodeficiency, the use of some types of exogenous interferons (e.g. Interferon-gamma) as immunomodulators has been attempted with therapeutic purposes, aiming at improving the TH-1 type immune response in HIV-infected and non-HIV-infected patients with tuberculosis.
The justification for the use of exogenous Interferon-gamma in these patients is due to the fact that the referred cytokine is known for its ability to stimulate a TH-1 type immune response, leading to macrophage reactivation, to increase, or at least partially recover, the ability of host immune mechanisms to respond to pathogens.
Animal Models Used for Studies of Intracellular Parasite Infectious Listeria Monocytogenes—Use of Immunomodulator
Among the several classical experimental models for the study of mechanisms of infections caused by strict or facultative intracellular parasites it is worth mentioning the model based on infection in experimental animals by the bacterium Listeria monocytogenes (L. monocytogenes), a pathogenic intracellular microorganism that like all other strict or facultative intracellular parasites counts on a survival and infection mechanism in the host body based on its capacity to take advantage of defective immune response or immunological deficiency.
This well-known infection model has been used for decades in the study of intracellular bacterial infection and in studies of cell-mediated immunity following these infections. (Tripathy S. P. & Mackaness G. B. 1969, the effect of cytotoxic agents on the primary response to Listeria monocytogenes. J. Exp. Med. 130:1-16).
For these reasons, the experimental model of L. monocytogenes infection is also appropriate for the study and development of new combinations of substances or medications, especially for medications or substances capable of influencing the host's immune response to microbial infections.
Thus, the medications or substances used in the treatment of infections caused by L. monocytogenes, including those with immune modulatory activity and found to be successful in these experiments may also be considered for use in the treatment of infections caused by intracellular pathogens of other genera and species, because their infection mechanisms are similar to those of the L. monocytogenes. 
In other words, substances with immunomodulatory activity which are efficient against the L. monocytogenes may be considered for use in preventive or curative therapies to be used in other types of intracellular parasites, such as M. tuberculosis, M. avium, M. leprae and Plasmodium sp, Leishmania sp, opportunistic fungi, as those of the Candida species, and finally in viral infections.
All these types of parasites are equally dependent on the existence or persistency of a status of immunological dysfunction, which, in turn, can be associated to a defective TH-1 type immune response of the host body to pathogen invasion.
This condition can be previous, concomitant or subsequent to the presence of microorganisms, being agreed in the present state of the art that it is necessary and/or facilitates the survival and dissemination of pathogens in the host body.
Therefore, the medications or therapies aimed at providing a more efficient immune response will be potentially useful in the treatment of diseases caused by a wide range of genera and species of microorganisms, including the above mentioned.
This was precisely the starting point for the selection of an immunomodulator (proteic aggregate of ammonium and magnesium phospholinoleate-palmitoleate anhydride) as one of the components of the combination of substances of the present invention.
State of the Art in the Treatment of Tuberculosis—Use of Drugs Specifically Target Against Mycobacterium Tuberculosis 
The main drugs specifically targeted against the parasites that cause tuberculosis (M. tuberculosis) known until now can be divided into four groups, according to their action in the several components of mycobacteria, as follows:
Group 1—bacterial cell wall inhibitors (eg. Isoniazid, Ethambutol, Ethionamide, Cycloserine); Group 2—bacterial nucleic acid synthesis inhibitors. (eg. Rifampicin, Quinolones); Group 3—bacterial protein synthesis inhibitors, (eg. Streptomycin, Kanamycin); Group 4—bacterial energy metabolism inhibitors (eg. Pirazinamide or PZA).
The antibacterial drugs or combinations of drugs developed until now to fight the mycobacteria that cause tuberculosis are able to target these bacteria in active growth or multiplication processes specifically, mostly by inhibiting bacterial cell processes, as shown above, and interfering with cell wall biogenesis and bacterial DNA replication. processes.
This means that the current drugs provide strong antibacterial action but low sterilizing activity, the latter being the property of eliminating or incapacitating the ability of bacteria in low growth phase and/or reduced metabolic activity to cause infection, which typically occurs in the intracellular life cycles of the M. tuberculosis and other mycobacteria.
Due to the referred inefficacy of current therapies to fight mycobacteria, in general, and the M. tuberculosis, in particular, and to the ability of these bacteria in deceiving the immune system and/or taking advantage of situations of host immune deficiency, these pathogens persist in the host organism even after treatment with association of Isoniazid (INH) with quinolones and fluoroquinolones (e.g. Levofloxacin, Moxifloxacin) and/or Pirazinamide (PZA), and even in case of associations of these drugs with injectable forms of other antibacterials, such as Kanamycin, Amikacin or Capreomycin.
State of the Art in Tuberculosis Treatment—Adjuvant Immunotherapy and the Benefits of Using the Present Invention
The biological characteristics of infection by M. tuberculosis, combined with the low efficacy of antituberculosis treatments that use the above mentioned drugs make it difficult to eradicate this type of microorganism.
The persistency of latent or hidden forms of the M. tuberculosis inside host immune system cells where they are not reached by current medication, indicates that the disease in its clinical forms may relapse and become a devastating event many years after the primary infection or the end of treatments, leading to the occurrence of tuberculosis in its typical clinical characteristics, by taking advantage of a situation of disease or defective immunity, such as in the case of Diabetes mellitus or an infection by HIV virus or AIDS.
Therefore, the expansion of AIDS epidemic, particularly in developing countries, not surprising, led to an explosive increase in the number of tuberculosis cases, which is related to the emergence of varieties of bacteria that become more resistant to treatment. Immunotherapy is, therefore, a promising treatment against this disease which aims at recovering the immune defense mechanisms of the host, particularly the TH-1 type immune response, which enables the immune system to respond to mycobacteria.
Several state of the art medical literature reports indicate that the referred strategy, that is, immunotherapy may have beneficial effects in the clinical treatment of tuberculosis, through the use of exogenous interferons, such as Interferon-alpha and Interferon-gamma (A. Yola et al., Retrovirology 2006; 3 (Suppl 1): S38), Suarez-Mendez et al. BMC Infect. Dis. (2004; 4: 44) as therapeutic adjuvant.
Despite advancements in the treatment of tuberculosis which were made possible with immunotherapy as an adjuvant therapy, using exogenous interferons, this modality of treatment is plagued by considerable problems, as explained in the present report such as the occurrence of undesirable side effects and high treatment cost.
Since the success or failure of host colonization by M. tuberculosis, as by all other facultative or strict intracellular pathogens, is highly dependent of the type and/or efficacy of the immunological response of the host's body, the use of present invention, by the ability of one of the components, a specific immunomodulator, to induce a potent TH-1 type response in the host, will undoubtedly make available a new and powerful weapon against that infectious microorganisms.
Additionally, the use of the combination of substances of the present invention against infections caused by mycobacteria, particularly the M. tuberculosis, will prevent or minimize the occurrence of major side effects, which are common in combinations of substances described in the state of the art that depend on exogenous interferons.